Recently, an analytical method for studying apparently invisible biological phenomena by determining metabolites contained in organisms comprehensively and quantitatively is attracting attentions. The analytical method, which is also called metabolome analysis, is attempted to be applied to the medical field, for example in disease diagnosis and etiological analysis, and also to the pharmaceutical field, for example in analysis of toxicity or adverse reaction. In addition to the medical and pharmaceutical fields, the analytical method is expected to be applied to other fields, for example for food quality control, quality rating, quality prediction, food stability evaluation, optimization of food production and breeding of industrial microbes, plants and others.
As described above in the metabolome analysis, there exists a need for conducting a quantitative analysis for quantitative determination of the substances to be analyzed. A gas chromatography mass spectrometer or a liquid chromatography mass spectrometer is generally used as such an instrument for quantitative analysis. However, substances to be analyzed such as biological metabolites are highly water-soluble low-molecular weight compounds such as amino acids, organic acids, and glucides, so that these compounds need to be derivatized for analysis on these analyzers. Thus, for comprehensive and quantitative analysis of analyte substances contained in the test sample, it is needed first to obtain many analyte substances from a collected test sample comprehensively, derivatize these substances and thus prepare a test sample solution suited for these analyzers.
Traditionally, for preparation of such a test sample solution, a sample was dried for complete removal of water; the dried sample was mixed with a prescribed substance sufficiently and heated for 60 to 90 minutes for modification of the carbonyl, amino, and hydroxyl groups of amino acids and glucides; further, a derivatization reagent was added thereto and the mixture was agitated sufficiently and heated for 30 to 60 minutes; and the resulting solution was analyzed within 24 hours after its preparation. In this way, there were needed a significant period of time and much labor for preparation of a test sample for metabolome analysis, and thus metabolome analysis was limited in applicability, although its usefulness was attracting attentions.
As a method to overcome the problem above, Patent Document 1 discloses a kit employing a pipette that can make free amino acids contained in sample be adsorbed on an ion-exchange resin by treatment of a test sample with the ion-exchange resin, elute the amino acids therefrom with an ion-exchanging elution medium, i.e., water or an aqueous salt solution, and derivatize (alkylate or esterify) the free amino acids contained in the eluate. It is also described that the free amino acids are eluted with an elution medium, liberated and alkylated or esterified in the presence of the elution medium and the ion-exchange resin, if it is a styrene-divinylbenzene copolymer-based ion-exchange resin.
Alternatively, Patent Document 2 discloses a pre-analysis treatment device that can separate analyte substances from a test sample so as to obtain a test sample solution.
However, although Patent Document 1 discloses a kit employing the pipette, it does not disclose a device that can be automated to collect the derivatized amino acids. In addition, derivatization of amino acids described therein was alkylation or esterification and there was no description on trimethylsilylation. Although separation of some analyte substances such as residual agricultural chemicals and environmental hormones, contained in the test sample was described in Patent Document 2, there was no description on derivatization of the analyte substances for subsequent analysis or on the instrument and the method that can give derivatized amino acids, organic acids and glucides.